Biotin (Vitamin H) is an important biomolecule that has been widely used in Biotinylation reactions to label antibodies or other proteins of interest. Due to its low molecular weight, biotin usually does not cause any significant changes in protein conformation and biological activity. Biotinylated proteins/antibodies are widely isolated and assayed with streptavidin pull-down assay, affinity chromatography, ELISA and Western blotting etc. Biovision offers a variety of ready-to-use Biotinylation reagents (Cat. # 2346, 2345, 2347, 2322, 2326, 2323). It is often desired to determine the degree of substitution of such biotinylated biomolecules. Biovision’s Biotin Assay Kit is based on the differential binding of Streptavidin to Biotin and a dye, 2-(4-hydroxyazobenzene) benzoic acid (HABA). When HABA is bound to Streptavidin, addition of free biotin or biotinylated biomolecules results in its displacement in the reaction. These changes are manifested in terms of decrease in the overall absorption (OD 500 nm) and can be quantitatively correlated to the amount of biotin present in solution. The kit also provides Biotin as a Standard and Biotinylated BSA as a positive control. This kit can be used to estimate as low as 60 pmol or 15 ng of biotin in solution.
|Cat # +Size||K811-100|
|Detection Method||Absorbance (500 nm)|
|Applications||Measuring free biotin or the substitution degree of Biotinylation (biotin labels) of proteins and antibodies|
|Features & Benefits||• This kit can be used to estimate as low as 60 pmol or 15 ng of biotin in solution|
|Kit Components||• Biotin Assay Buffer • Biotin Assay Reagent A • Biotin Assay Reagent B • Biotin Standard Solution (10 mM in DMSO) • Biotinylated BSA (2 mg/ml)|
|Shipping Conditions||Gel Pack|
|USAGE||For Research Use Only! Not For Use in Humans.|
What type of trypsin should be used with the samples?
Any trypsin is fine as long as its activity is good. We recommend using 1-5%, which is the highest concentration. It could be lower if the trypsin activity is high. EDTA should be OK as well and it does not interfere the Biotin quantification. What concentration of trypsin is recommended for the protein digestion in step VI
according to K811-100 protocol? Is EDTA compatible with the kit?
Any trypsin is fine as long as it has good activity. We recommended 1-5%, which is the highest concentration that can be used. It could be lower if the trypsin activity is very good. EDTA should be OK and it does not interfere with the Biotin assay.
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